Biology subjects

Lee, C. P.

Publications and source records attributed to Lee, C. P..

4 recordsLinked to original sources

bioRxiv2021

Enzymes degraded under high light maintain proteostasis by transcriptional regulation in Arabidopsis

Photo-inhibitory high light stress in Arabidopsis leads to increases in markers of protein degradation and transcriptional upregulation of proteases and proteolytic machinery, but proteostasis is largely maintained. We find significant increases in the in vivo degradation rate for specific molecular chaperones, nitrate reductase, glyceraldehyde-3 phosphate dehydrogenase, and phosphoglycerate kinase and other plastid, mitochondrial, peroxisomal, and cytosolic enzymes involved in redox shuttles. Coupled analysis of protein degradation rates, mRNA levels, and protein abundance reveal that 57% of the nuclear-encoded enzymes with higher degradation rates also had high light-induced transcriptional responses to maintain proteostasis. In contrast, plastid-encoded proteins with enhanced degradation rates showed decreased transcript abundances and must maintain protein abundance by other processes. This analysis reveals a light-induced transcriptional program for nuclear-encoded genes, beyond the regulation of PSII D1 subunit and the function of PSII, to replace key protein degradation targets in plants and ensure proteostasis under high light stress.

plant biology↗

bioRxiv2021

Defects in autophagy lead to selective in vivo changes in turnover of cytosolic and organelle proteins in Arabidopsis

Identification of autophagic protein cargo in plants by their abundance in autophagy related genes (ATG) mutants is complicated by changes in both protein synthesis and protein degradation. To detect autophagic cargo, we measured protein degradation rate in shoots and roots of Arabidopsis atg5 and atg11 mutant plants. These data show that less than a quarter of proteins changing in abundance are probable cargo and revealed roles of ATG11 and ATG5 in degradation of specific cytosol, chloroplast and ER-resident proteins, and a specialized role for ATG11 in degradation of proteins from mitochondria and chloroplasts. Our data support a role for autophagy in degrading glycolytic enzymes and the chaperonin containing T-complex polypeptide-1 complex. Autophagy induction by Pi limitation changed metabolic profiles and the protein synthesis and degradation rates of atg5 and atg11 plants. A general decrease in the abundance of amino acids and increase in several secondary metabolites in autophagy mutants was consistent with altered catabolism and changes in energy conversion caused by reduced degradation rate of specific proteins. Combining measures of changes in protein abundance and degradation rates, we also identify ATG11 and ATG5 associated protein cargo of low Pi induced autophagy in chloroplasts and ER-resident proteins involved in secondary metabolism. Single Sentence SummaryProtein cargo of autophagy in plants can be discovered by identifying proteins that increase in abundance and decrease in degradation rate in mutants deficient in autophagy machinery

plant biology↗

bioRxiv2021

The mitochondrial pyruvate carrier (MPC) complex is one of three pyruvate-supplying pathways that sustain Arabidopsis respiratory metabolism

Malate oxidation by plant mitochondria enables the generation of both oxaloacetate (OAA) and pyruvate for tricarboxylic acid (TCA) cycle function, potentially eliminating the need for pyruvate transport into mitochondria in plants. Here we show that the absence of the mitochondrial pyruvate carrier 1 (MPC1) causes the co-commitment loss of its orthologs, MPC3/MPC4, and eliminates pyruvate transport into Arabidopsis mitochondria, proving it is essential for MPC complex function. While the loss of either MPC or mitochondrial pyruvate-generating NAD-malic enzyme (NAD-ME) did not cause vegetative phenotypes, the lack of both reduced plant growth and caused an increase in cellular pyruvate levels, indicating a block in respiratory metabolism, and elevated the levels of branched-chain amino acids at night, a sign of alterative substrate provision for respiration. 13C-pyruvate feeding of leaves lacking MPC showed metabolic homeostasis were largely maintained except for alanine and glutamate, indicating that transamination contributes to restoration of the metabolic network to an operating equilibrium by delivering pyruvate independently of MPC into the matrix. Inhibition of alanine aminotransferases (AlaAT) when MPC1 is absent resulted in extremely retarded phenotypes in Arabidopsis, suggesting all pyruvate-supplying enzymes work synergistically to support the TCA cycle for sustained plant growth.

plant biology↗

bioRxiv2020

The Arabidopsis mitochondrial dicarboxylate carrier 2 maintains leaf metabolic homeostasis by uniting malate import and citrate export

Malate is the major substrate for respiratory oxidative phosphorylation in illuminated leaves. In the mitochondria malate is converted to citrate either for replenishing tricarboxylic acid (TCA) cycle with carbon, or to be exported as substrate for cytosolic biosynthetic pathways or for storage in the vacuole. In this study, we show that DIC2 functions as a mitochondrial malate/citrate carrier in vivo in Arabidopsis. DIC2 knockout (dic2-1) results in growth retardation that can only be restored by expressing DIC2 but not its closest homologs DIC1 or DIC3, indicating that their substrate preferences are not identical. Malate uptake by non-energised dic2-1 mitochondria is reduced but can be restored in fully energised mitochondria by altering fumarate and pyruvate/oxaloacetate transport. A reduced citrate export but an increased citrate accumulation in substrate-fed, energised dic2-1 mitochondria suggest that DIC2 facilitates the export of citrate from the matrix. Consistent with this, metabolic defects in response to a sudden dark shift or prolonged darkness could be observed in dic2-1 leaves, including altered malate, citrate and 2-oxoglutarate utilisation. There was no alteration in TCA cycle metabolite pools and NAD redox state at night; however, isotopic glucose tracing reveals a reduction in citrate labelling in dic2-1 which resulted in a diversion of flux towards glutamine, as well as the removal of excess malate via asparagine and threonine synthesis. Overall, these observations indicate that DIC2 is responsible in vivo for mitochondrial malate import and citrate export which coordinate carbon metabolism between the mitochondrial matrix and the other cell compartments. SIGNIFICANCE STATEMENTMitochondria are pivotal for plant metabolism. One of their central functions is to provide carbon intermediates for the synthesis of critical building blocks, such as amino acids. Malate import and citrate export are two of the most recognised and specialised features of the mitochondrial role in the plant cellular metabolic network, yet the possibility that a single carrier would unite both functions has not been considered. Here, we have demonstrated that DIC2 preferentially fulfils these two functions in Arabidopsis thaliana in vivo, making it a bifunctional gateway for two major metabolite fluxes into and out of the mitochondrial matrix in the plant cell. Our results highlight the significance of DIC2 in cooperation with other mitochondrial carriers in maintaining metabolic balance even under challenging environmental conditions.

plant biology↗